No abstract

No abstract

No abstract

Hydrogen peroxide, one of the potent oxidants in muscle tissues, can induce very rapid oxidation of oxymyoglobin (MbO2) to metmyoglobin (metMb) with an apparent rate constant of 7.5 X 10(4) h-1 M-1 (i.e., 20.8 s-1 M-1) over the wide pH range of 5.5-10.2 in 0.1 M buffer at 25 degrees C. Its molecular mechanism, however, is quite different from that of the autoxidation of MbO2 to metMb. Kinetic analysis has revealed that the hydrogen peroxide oxidation proceeds through the formation of ferryl-Mb(IV) from deoxy-Mb(II), which is in equilibrium with MbO2, by a two-equivalent oxidation with H2O2. Once the ferryl species is formed, it reacts rapidly with another deoxy-Mb(II) in a bimolecular fashion so as to yield 2 mol of metMb(III). Under physiological conditions, the rate-determining step was the oxidation of the deoxy species by H2O2, its rate constant being estimated to be on the order of 3.6 X 10(3) s-1 M-1 at 25 degrees C. These findings leads us to the view that a good supply of dioxygen provides rather an important defense against the oxidation of myoglobin with hydrogen peroxide in cardiac and skeletal muscle tissues.

Human oxyhemoglobin showed a biphasic autoxidation curve containing two rate constants, i.e. k f for the fast autoxidation due to the ␣ chains, and k s for the slow autoxidation of the ␤ chains, respectively. Consequently, the autoxidation of the HbO 2 tetramer produces two different curves from the pH dependence of k f and k s . The analysis of these curves revealed that the ␤ chain of the HbO 2 tetramer does not exhibit any protoncatalyzed autoxidation, unlike the ␣ chain, where a proton-catalyzed process involving the distal histidine residue can play a dominant role in the autoxidation rate. When the ␣ and ␤ chains were separated from the HbO 2 tetramer, however, each chain was oxidized much more rapidly than in the tetrameric parent. Moreover, the separated ␤ chain was recovered completely to strong acid catalysis in its autoxidation rate. These new findings lead us to conclude that the formation of the ␣ 1 ␤ 1 contact produces in the ␤ chain a conformational constraint whereby the distal histidine at position 63 is tilted away slightly from the bound dioxygen, preventing the proton-catalyzed displacement of O 2. by a solvent water molecule. The ␤ chains have thus acquired a delayed autoxidation in the HbO 2 tetramer.The reversible and stable binding of molecular oxygen to the heme iron(II) is the basis of hemoglobin function. Consequently, much attention has been directed to the elucidation of the molecular mechanism of cooperative oxygen binding to the hemoglobin tetramer (1). However, the oxygenated form of hemoglobin, as well as of myoglobin, is known to be oxidized easily to the ferric(III) met form, which cannot bind molecular oxygen and is therefore physiologically inactive, with generation of the superoxide anion (2-5).To this autoxidation reaction, it has been widely accepted that hemoglobin is much more resistant as compared with myoglobin. Unlike myoglobin, Mansouri and Winterhalter (6) reported that oxyhemoglobin (HbO 2 ) showed a biphasic autoxidation reaction with a fast and a slow component. They also demonstrated that the ␣ chain was oxidized more rapidly than the ␤ chain in hemoglobin tetramer. At the same time, however, there have been a number of reports that such a rate difference was not observed between the ␣ and ␤ chains in the autoxidation reaction of HbA (7-9). Rather, Zhang et al. (9) showed that the rate of autoxidation was markedly enhanced when the HbO 2 tetramer dissociates into ␣␤ dimers. To clarify these discrepancies, we have recently examined systematically the effect of hemoglobin concentration on the autoxidation rate at several different values of pH, and found that human HbO 2 exhibits a biphasic autoxidation curve only in the pH range from neutral to acidic (5). By dissociation of tetramers into ␣␤ dimers, the rate of autoxidation for the fast component (due to the ␣ chain) was also found to increase markedly at the acidic pH, but the addition of 2,3-diphosphoglyceric acid offered no significant effect on the increment of the autoxidation rate (5).In the present pape...